The MALDI-MS technique, or Matrix-Assisted Laser Desorption Ionization Mass Spectrometry [M. Karas, F Hillencamp., Anal. Chem., 60, 2301 (1988); R. C. Beavis., Org. Mass. Spec., 27, 653 (1992)], has emerged as a powerful analytical tool in the investigation of non-volatile high molecular weight compounds, especially fragile biological molecules. Such techniques are simply referred to as “MALDI” herein. In the matrix-assisted laser desorption process, the anaylate is mixed with a matrix, usually a small organic molecule, at mole ratios of few thousand matrix molecules to the analyte. During irradiation with a laser, the matrix molecules assist in the desorption of the analyte molecule as molecular ions, as shown in FIG. 1. The mechanistic aspects of the interaction of the matrix molecules and the anaylate during irradiation has been a topic of continuing research [R. Zenobi, R. Knochenmuss, Mass. Spec. Rev., 17, 337 (1998)]. MALDI is a soft-ionization technique, where the energy from the incident laser radiation is utilized to desorb the small molecules rather than dissociating the anaylate.
One of the key issues in MALDI analysis is the selection and availability of the right matrix molecule. The matrix molecule, in its assisting role, is expected to function as an efficient absorber of laser energy—as well to isolate the polymer molecules from one another [M. Karas, F. Hillencamp, R. C. Beavis, B. T. Chait., Anal. Chem., 63, 1193A (1991); H. S. Creel., Trends. Pol. Sci., 1, 336 (1993)]. While the purpose of the matrix is to desorb the anaylate without degradation, there is a positive aspect to fragmentation, especially when the anaylate is a biomolecule such as a peptide, protein or DNA.
Tandem mass spectrometry (TMS), in the recent past, has been developed into an efficient technique for solving the structural problems and sequencing of proteins and DNA molecules [W. D. Bowers, S. S. Delbert, R. L. Hunter, R. T. McIver., J. Am. Chem. Soc, 106, 7288 (1984); K. Biemann et al., in Mass Spectrometry in the Analysis of Large Molecules, C. J. McNeal ed (Wiley, Chichester, 1986)]. The schematics of tandem mass spectrometry are shown in FIG. 2. Referring to FIG. 2, a combination of biomolecules are ionized by fast ion bombardment at stage A, after which they pass through the first mass selector (M1). The mass selected species then passes to the collision chamber (CS) where fragmentation is achieved through collision-induced decomposition (CID). The mass distribution of the fragments is analyzed in a second mass spectrometer (M2) and a detector (D) to generate a CID-MS spectrum. The applications of tandem mass spectrometry in the sequencing of biomolecules has recently been reviewed by Biemann and Scoble [K. Biemann, H. A. Scoble., Science, 237, 992 (1987)].
A look at the fundamentals of both these techniques reveal that a matrix with properties tunable from one extreme of complete analyte desorption to the other extreme of complete fragmentation of the analyte can combine the advantages of both MALDI and TMS in a single technique. This requires a unique matrix element that is an efficient absorber in the ultraviolet (UV) region of the electromagnetic (EM) spectrum, and which can go through photo-ionization followed by excited state proton transfer (ESPT) [M. Karas, D. Bachmann, U. Bahr, F. Hillenkamp., Int. J. Mass. Spectrm. Ion. Proc., 78, 53 (1987)]. The ESPT process requires the presence of labile protons in the matrix molecule. Some of the traditional matrix molecules are shown in FIG. 3. A common characteristic of all these matrix molecules is the presence of a labile, acidic proton in their molecular structure.